The overall objective of this research program has been to identify and characterize the actions of hormone (LH) and prolactin (PRL) from the anterior pituitary gland in response to the central neurotransmitter and neuropeptide systems that regulate the secretion of luteinizing physiological stimuli. Neuropeptide Y (NPY) functions in this system as an amplifier of the signals provided by the adrenergic transmitters to stimulate LH-releasing hormone (LHRH) secretion and also augments the signal provided to the gonadotrophe by LHRH. In this application, we propose to extend this research into a new physiological context, following the recent demonstrations that the synthesis of NPY in the medial-basal hypothalamus is up-regulated in lactation, and that at least part of this is attributable to a novel expression of the peptide in the tuberoinfundibular dopamine (TIDA) neurons. At present, neither the biological significance of, nor the physiological signals that evoke, this state-specific altered expression of NPY are known, and the proposed studies are designed to address these two questions. Because the ADA system is the major neuroendocrine PRL-inhibiting system, we hypothesize that the increased NPY present in the medial basal hypothalamus may affect PRL secretion from the anterior pituitary gland by modulating the action of DA on the lactotrophes and/or the release of DA from the hypothalamus. Second, we will examine whether the suppression of gonadotropin secretion during lactation that is produced by the suckling stimulus and elevated PRL is mediated by the increased NPY via an interaction with endogenous opioids. As the third main objective, we will test whether PRL and/or the suckling stimulus is the physiological signal that leads to increased NPY expression in the medial-basal hypothalamus during lactation. The first Specific Aim contains in vitro studies that will investigate the effects of NPY and NPY-DA interactions on second messenger systems and PRL secretion in cultured anterior pituitary cells. The second Specific Aim will use in vitro and in vivo approaches to examine whether NPY modulates the release of DA from the median eminence of lactating rats. In vivo studies of the third Specific Aim will determine the roles of DA and NPY in generating the episodic pattern of PRL secretion during lactation and measure the pattern of NPY and DA release during nursing. The fourth Specific Aim will investigate whether an NPY-endogenous opioid interaction mediates the inhibition of gonadotropin secretion during lactation by testing the in vivo effects of NPY and NPY antagonists, and by examining NPY-opioid interaction, in episodic LH secretion and LHRH receptor regulation in lactating rats. These studies will also investigate in vitro effects of NPY on LHRH and Beta-endorphin release from medial-basal hypothalamus of lactating rats. The fifth Specific Aim will evaluate whether the suckling stimulus and/or a central action of PRL is the physiological signal responsible for enhanced NPY expression in the medial basal hypothalamus during lactation. These studies will test the effects of producing hyperprolactinemia in non-lactating rats and of suppression of PRL release in lactating rats on a) NPY immunoreactivity in the medial basal hypothalamus (by RIA), b) NPY immunoreactivity and innervation pattern in TIDA and non-TIDA neurons (by light and EM immunocytochemistry), and c) preproNPY mRNA levels in TIDA and non-TIDA neurons (by in situ hybridization).